Ammonia Excretion Rates Research Articles

Gene knockout of NHX-3 in the soil nematode Caenorhabditis elegans leads to broad-spectrum compensatory regulation of Na+/H+ exchangers, antiporters, and the V-type H+-ATPase

Na+/H+ exchangers are directly involved in a variety of an animal's essential physiological processes such as ionoregulation, acid-base regulation, nitrogenous waste excretion, and nutrient absorption. While nine NHX isoforms have been identified in Caenorhabditis elegans, the physiological importance of each isoform is not understood. The current study aimed to further our knowledge of NHX-3 which has previously been suggested to be involved in the movement of ammonia and acid-base equivalents across the nematode's hypodermis. Although NHX-3 knockout mutant nematodes exported H+ and imported Na+ at slower rates than wild-type nematodes, attempts to inhibit the NHX activity of mutant nematodes using amiloride and EIPA caused an unexpected increase in hypodermal H+ export and did not impact Na+ fluxes suggesting that the different H+ and Na+ transport profiles of the nematodes are likely due to compensatory changes in the mutants in response to the NHX-3 knockout, rather than the loss of NHX-3's physiological function. Significant changes in the mRNA expression of 7 other NHX isoforms, 2 Na+/H+ antiporter isoforms, and the V-type H+-ATPase were detected between wild-type and mutant nematodes. Furthermore, mutant nematodes possessed significantly reduced rates of cytochrome C oxidase activity and ammonia excretion rates, indicating the knockout of NHX-3 induced fundamental changes in metabolism that could impact the nematode's need to eliminate metabolic end-products like H+ and ammonia that relate to NHX transport. While C. elegans is a popular genetic model with cheap and accessible commercial mutants, our findings suggest caution in interpretation of results in studies using mutants to study physiological traits and the biological significance of specific transporters.

The effects of inbreeding on stress resistance of the Pacific oyster Crassostrea gigas at different temperatures and salinities

ABSTRACT The Pacific oyster (Crassostrea gigas) is a commercially important shellfish widely cultured worldwide. Understanding the effect of inbreeding on C. gigas is critical to the long-term feasibility of breeding programmes, especially when selected lines are developed in hatcheries with limited effective population sizes. The effect of inbreeding on stress resistance in C. gigas remains to be explored. The present study evaluated the physiological and immune responses to different temperatures (16–36°C) and salinities (20–40 psu) in an inbreeding line and a wild population of C. gigas. Two physiological parameters, including ammonia-N excretion rate (AER) and oxygen consumption rate (OCR), and three enzyme activities including superoxide dismutase activity (SOD), catalase activity (CAT), and contents of malondialdehyde (MDA) were measured on day 14 of the temperature and salinity exposure. Compared with the wild population, the physiological parameters (AER and OCR) were significantly lower, and the enzyme activities (SOD, CAT, and MDA) were significantly higher in the inbreeding line at suboptimal temperatures or salinities. These results showed that inbreeding has negative effects on stress resistance in C. gigas. In addition, multiple groups with different inbreeding levels would be needed to quantify the effects of inbreeding on stress resistance in C. gigas.

Energetic budget of diploid and triploid eastern oysters during a summer die-off.

Triploid oysters are widely used in off-bottom aquaculture of eastern oysters, Crassostrea virginica. However, farmers of the Gulf of Mexico (GoM) and Atlantic coast estuaries have observed unresolved, late-spring die-offs of triploid oysters, threatening the sustainability of triploid aquaculture. To investigate this, the physiological processes underlying oyster growth (e.g., feeding, respiration) and mortality of one-year-old diploid and triploid oysters were compared in early summer following an uptick in mortality. It was predicted that higher triploid mortality was the result of energetic imbalances (increased metabolic demands and decreased feeding behavior). Oyster clearance rates, percentage of time valves were open, absorption efficiency, oxygen consumption rates (basal and routine), ammonia excretion rate were measured in the laboratory and scope for growth was calculated. In addition, their condition index, gametogenic stage, Perkinsus marinus infection level, and mortality were measured. Mortality of triploids in the laboratory was greater than for diploids, mirroring mortality observed in a related field study. The physiological parameters measured, however, could not explain triploid mortality. Scope for growth, condition index, and clearance rates of triploids were greater than for diploids, suggesting sufficient energy reserves, while all other measurements where similar between the ploidies. It remains to be determined whether mortality could be caused from disruption of energy homeostasis at the cellular level.

Filtration capabilities of freshwater mussel (Lamellidens marginalis) and apple snail (Pila globosa) and their potential impacts on freshwater integrated multitrophic aquaculture systems

Aim: The study aimed to assess the relative filtration rates of an indigenous freshwater mussel, Lamellidens marginalis and an indigenous freshwater snail, Pila globosa, and their suitability for incorporation into freshwater integrated multi-trophic aquaculture (FIMTA) systems. Methodology: An indoor experimental trial was conducted in a wet laboratory (temperature: 30.5 ± 0.5, light intensity: 2000 lux, light cycle: L: D: 12hr: 12hr). The trial was performed in glass aquaria in triplicate filled with algae rich water to assess their filtration rate. Simultaneously, the impacts of these two molluscs on inorganic nitrogen and phosphorus were evaluated. Results: L. marginalis exhibited filtration rates of 934.7± 0.07 cells min-1 mussel-1 and 13.79 ± 0.04 cells g-1 min-1) against 261.8 ± 10.15 cells min-1 snail-1 and 9.74±0.15 cells g-1 min-1 for P globosa. Similarly, the mussel also exhibited markedly higher turbidity and chlorophyll clearance capacity (p<0.05) in comparison to snail, especially during initial 6-12 hr when initial concentrations of algal population, turbidity and Chl-a were high. Notably, the oxygen consumption and ammonia excretion rates were greater for snail than mussel after 24hr and at the end of the experiment. Interpretation: The results provide quantitative information on the filtration capacity of two indigenous freshwater molluscs that have considerable local food as well as economic value and would pave the way for their utilization as ecological engineers, especially organic extractive in freshwater integrated multi-trophic aquaculture (FIMTA) system. It is concluded from the study that L. marginalis was more efficient in extracting almost four times algal cells than, P. globosa. Key words: Eutrophication, Filtration, Lamellidens marginalis, Pila globosa

Validation of target protein PcnWAS in Pomacea canaliculata and screening of target-based molluscicidal compounds

Virtual screening is an efficient way to obtain new drugs, which has become an important method in the field of pesticide research. Protein neural wiskott-Aldrich syndrome isoform X1 (PcnWAS) is a target protein that exists in the haemocytes of Pomacea canaliculata, and in this study, isothermal titration calorimetry (ITC) was used to evaluate the binding ability of protein PcnWAS and pedunsaponin A in vitro. Furthermore, it was set as a receptor, and the design of molluscicidal compounds based on protein PcnWAS was carried out. Results showed that, pedunsaponin A had high binding capacity with protein PcnWAS, and the binding constant (Ka) was 2.98 ± 1.74 × 10−4. A new potential molluscicidal compound thionicotinamide-adenine-dinucleotide (thionicotinamide-DPN) was obtained by virtual screening. In-vivo bioassay indicated that, the LC50 value was 57.7102 mg/L (72 h), and the oxygen consumption rate, ammonia excretion rate, oxygen nitrogen ratio and hemocyanin content of P. canaliculata declined after 60 mg/L thionicotinamide-DPN treated. Furthermore, the treatment of thionicotinamide-DPN also decreased gene expression level of protein PcnWAS. The results of ITC test showed that thionicotinamide-DPN can bind with protein PcnWAS efficiently, which means that it has the same target with pedunsaponin A when interacted with P. canaliculata. All the above results lay a foundation for the development of new molluscicides.

Growth performance and quality indicators of European seabass (Dicentrarchus labrax) fed diets including refined glycerol

Glycerol is a by-product of the biodiesel industry with growing availability and applicability in animal feeds. However, its use as a source of energy in aquafeeds has only been investigated in a small number of fish species. The effects of dietary refined glycerol inclusion rate on growth performance, nitrogen and energy balances, and fish quality were investigated in juvenile European seabass. Twelve fish were assigned to each of the nine tanks and used to test the experimental diets in triplicate. The diets comprised 0, 25, or 50 g kg−1 of refined glycerol (G0, G25, and G50, respectively), being not isoenergetic, although having similar digestible protein-to-energy ratios (22 mg kJ−1). Fish were fed twice a day to apparent satiation for 90 days. The G25 group displayed a higher absolute feed intake and a higher weight gain than the G0 or G50 groups, although feed efficiency and the use of protein as an energy source were similar between the three groups. Seabass fed the G25 diet also displayed a higher specific growth rate than fish fed the G50 diet. Higher hepato-somatic and visceral lipid-somatic indices of seabass were observed with increased levels of dietary glycerol, although proximal body composition remained similar across groups. The apparent digestibility coefficients for dry matter, crude protein, and energy increased with increasing dietary glycerol inclusion rate, whereas the apparent lipid digestibility coefficient was highest for the G25 group. No differences in the energy retained as protein, nitrogen efficiency, and total ammonia excretion rate were detected between seabass fed the different diets. At the end of the growth trial, physical and microbiological quality parameters of the skin and gill surfaces were assessed after 1 and 10 days of storage on ice. Fish quality decreased with storage time but was not affected by the dietary glycerol inclusion rate. In summary, the inclusion of refined glycerol in seabass diets may have economic benefits for feed manufacture and a positive effect on growth performance but does not change the efficiency of protein utilization in this carnivorous species.

Investigation of physiological energetic response of the thick shell mussel, Mytilus coruscus, to microplastics and low salinity: Potential countermeasures to multi-environmental changes

Microplastics (MPs) contamination and salinity fluctuation are two common phenomena on the East China Sea coast. The purpose of this experiment was to evaluate the combined effects of MPs and salinity on the energy budget of the thick shell mussel Mytilus coruscus. Juvenile mussels were exposed to six combined treatments with three MPs levels (0, 10 and 1000 items L−1) and two salinity levels (15, 25) for 28 d. The clearance rate, food absorption efficiency, respiration rate, ammonium excretion rate, and O:N ratio were significantly reduced, and the fecal organic dry weight ratio was significantly increased at salinity 15, inducing a reduction in the scope for growth. Under salinity 15, 1000 items L−1 MPs had significant negative effects on growth at Day 28. However, under salinity 25, the results were opposite in that the level of 1000 items L−1 MPs brought an obvious increment in scope for growth at Day 28. This meant that there were interactions between salinity and MPs in scope for growth at Day 28. These results suggest that the physiological energetics of juvenile M. coruscus can acclimate to MPs contamination with little physiological effect under normal salinity. This experiment has revealed that although marine MPs contamination is of concern, marine organisms likely have a significant capacity to adapt to it.

Discovery of a Potential Molluscicide Based on Protein PcRoo in Gill Cilia of Pomacea canaliculata.

In modern pesticide discovery, target-based drug design is an attractive and cost-effective approach. Previous studies found that protein rootletin (PcRoo) is a target protein of arecoline, when interacted with Pomacea canaliculata. In this study, we modeled the target protein through threading, and the binding energy between arecoline and protein PcRoo was calculated as -5.02 kcal/mol by molecular docking. Furthermore, two target compounds, baclofen and acedoben, with molluscicidal activity in theory were obtained by virtual screening in database DrugBank. The in vivo bioassay showed that baclofen could induce typical poisoning symptoms on P. canaliculata, which were characterized by weakness of foot muscles and loss of gill cilia, and the LC50 value was 16.2437 mg/L (72 h). Additionally, after 15 mg/L baclofen treatment, the oxygen consumption rate, ammonia excretion rate, and oxygen nitrogen ratio of P. canaliculata declined. Furthermore, the treatment of baclofen also decreased the gene expression level of PcRoo. These trends were the same as the changes after 5 mg/L arecoline treatment. The pharmacophore characteristics were further analyzed, and the results showed that the chemical structures of baclofen and arecoline were correlated in molluscicidal activity. These findings indicate that baclofen has the potential to be used as a molluscicide in agricultural production, and other new molluscicides may be obtained by virtual screening based on protein PcRoo.

Physiological and biochemical responses of the estuarine pulmonate mud snail, Amphibola crenata, sub-chronically exposed to waterborne cadmium

Physiological and biochemical responses of the pulmonate mud snail, Amphibola crenata, to waterborne cadmium (Cd) were investigated to determine the mechanisms of toxicity and impacts of a 21-d Cd exposure. Mud snails were exposed to nominal Cd concentrations of 0, 0.2, 4 and 8 mg L − 1 and bioaccumulation, whole animal physiological (oxygen consumption, ammonia excretion and oxygen:nitrogen), and tissue level biochemical (catalase activity, lipid peroxidation, glycogen, glucose and protein) endpoints were measured every 7 days. At the two highest Cd exposure concentrations complete mortality was observed over 21-d. In surviving animals, oxygen consumption declined and ammonia excretion rate increased with Cd exposure concentration and duration. The increased ammonia excretion likely reflected enhanced protein metabolism as suggested by a reduced oxygen:nitrogen (O:N). Increasing waterborne Cd concentration and exposure time led to increasing metal accumulation in all tissues. The snail viscera showed the highest Cd accumulation. Both catalase activity and lipid peroxidation in the viscera significantly increased with Cd exposure concentration and time, whereas, the foot muscle and remaining tissues (kidney, mantle, remaining digestive tissues and heart) showed increased catalase activity and lipid peroxidation at higher Cd concentrations (4 and 8 mg L − 1), suggestive of an effect of Cd on oxidative stress. Over the course of 21 days, Cd exposure resulted in significantly lower levels of glycogen in viscera relative to Cd-free controls, reflecting an increased energy demand. Haemolymph glucose rose initially and then fell with increased exposure duration, while haemolymph protein generally exhibited an increased concentration in Cd-exposure groups, reflecting the changes in energy substrates noted for somatic tissues. These results suggest that the physiological and biochemical responses of A. crenata to Cd are conserved relative to other aquatic animals, and were tissue-specific, dose- and time-dependant.

Behavioural and physiological responses to salinization and air exposure during the ontogeny of a freshwater South American snail.

Salinization is of global concern, threatening freshwater biodiversity. Salinity tolerance is highly variable and therefore needs to be evaluated on a species-specific basis. An estuarine population of Chilina dombeiana, a freshwater gastropod endemic to Chile and classified as vulnerable, has been recently found in the Biobío River's mouth, suggesting some degree of tolerance to brackish waters. This study evaluated the survival, behaviour (medium preference) and physiology of C. dombeiana when exposed to salinities higher than freshwater, thus elucidating the potential mechanisms used to survive salinization. Chilina dombeiana belongs to the Pulmonate group;, so we evaluated oxygen uptake in air and water, aiming to evaluate emersion as a potential avoidance response to a progressive salinity increase. Complete embryo development was observed for salinities ≤ 16 PSU (practical salinity units) but hatching rates above 50% were only achieved in freshwater (0 PSU). It was also found that salinity had stage-specific effects during embryonic development. In adults, acute exposure to brackish water (12 PSU) caused a decrease in oxygen consumption (compared to freshwater), in the ammonium excretion rates and in the percentage of muscular water content. Although C. dombeiana was able to take up oxygen in both mediums, survival in air decreased over time (days), which correlates with the behavioural preference to remain submerged, even at elevated salinities. Considering the survival of adults and embryos decreased as salinity increased and the lack of an avoidance behaviour or a physiological ability to maintain homeostasis at salinities higher than freshwater, our results suggest this snail could be adversely affected by salinization in the long term. Furthermore, given the ability of C. dombeiana to uptake oxygen in both mediums, it should be considered as a facultative air breather snail, rather than a strictly aquatic species.

Exercise and emersion in airand recovery in seawater in the green crab (Carcinus maenas): Effects on nitrogenous wastes and branchial chamber fluid chemistry.

At low tide, the green crab, which is capable of breathing air, may leave the water and walk on the foreshore, carrying branchial chamber fluid (BCF). N-waste metabolism was examined in crabs at rest in seawater (32 ppt, 13°C), and during 18-h recovery in seawater after 1 h of exhaustive exercise (0.25 BL s-1 ) on a treadmill in air (20°C-23°C), or 1 h of quiet emersion in air. Measurements were made in parallel to O2 consumption (ṀO2 ), acid-base, cardio-respiratory, and ion data reported previously. At rest, the ammonia-N excretion rate (ṀAmm = 44 µmol-N kg-1 h-1 ) and ammonia quotient (AQ; ṀAmm /ṀO2 = 0.088) were low for a carnivore. Immediately after exercise and return to seawater, ṀAmm increased by 65-fold above control rates. After emersion alone and return to seawater, ṀAmm increased by 17-fold. These ammonia-N bursts were greater, but transient relative to longer-lasting elevations in ṀO2 , resulting in temporal disturbances of AQ. Intermittent excretion of urea-N and urate-N at rest and during recovery indicated the metabolic importance of these N-wastes. Hemolymph glutamate, glutamine, and PNH3 did not change. Hemolymph ammonia-N, urea-N, and urate-N concentrations increased after exercise and more moderately after emersion, with urate-N exhibiting the largest absolute increments, and urea-N the longest-lasting elevations. All three N-wastes were present in the BCF, with ammonia-N and PNH3 far above hemolymph levels even at rest. BCF volume declined by 34% postemersion and 77% postexercise, with little change in osmolality but large increases in ammonia-N concentrations. Neither rapid flushing of stored BCF nor clearance of hemolymph ammonia-N could explain the surges in ṀAmm after return to seawater.

Light spectrum impacts on development respiratory metabolism and antioxidant capacity of larval swimming crab Portunus trituberculatus

The effects of the spectrum on the development, respiratory metabolism, and antioxidant capacity of the larval swimming crab Portunus trituberculatus were studied. Seven light spectra, i.e., purple (400 nm), blue (425 nm), cyan (510 nm), green (525 nm), yellow (598 nm), red (638 nm), and white (full spectrum), were estimated. The larvae had the optimum survival rate and development under cyan light. On the contrary, larvae in red and yellow lights had poor growth performance. The oxygen consumption rate (OCR) dropped while the ammonia excretion rate (AER) rose as the larvae developed. Early larvae’s oxygen-nitrogen ratio (O: N) fell when exposed to red light, suggesting more protein was utilized in the respiratory process. Regarding the antioxidant system, crab had the lowest malondialdehyde (MDA) under green, cyan and yellow light, and the highest total antioxidant capacity (T-AOC) in cyan light. Taken together, the current results suggest that cyan was the optimum spectrum for the development of P. trituberculatus larvae.

Do global environmental drivers' ocean acidification and warming exacerbate the effects of oil pollution on the physiological energetics of Scylla serrata?

Global climate change-induced ocean warming and acidification have complex reverberations on the physiological functioning of marine ectotherms. The Sundarbans estuarine system has been under threat for the past few decades due to natural and anthropogenic disturbances. In recent years, petroleum products' transportation and their usage have increased manifold, which causes accidental oil spills. The mud crab (Scylla serrata) is one of the most commercially exploited species in the Sundarbans. The key objective of this study was to delineate whether rearing under global environmental drivers (ocean acidification and warming) exacerbates the effect of a local driver (oil pollution) on the physiological energetics of mud crab (Scylla serrata) from the Sundarbans estuarine system. Animals were reared separately for 30days under (a) the current climatic scenario (pH 8.1, 28°C) and (b) the predicted climate change scenario (pH 7.7, 34°C). After rearing for 30days, 50% of the animals from each treatment were exposed to 5mg L-1 of marine diesel oil for the next 24h. Physiological energetics (ingestion rate, absorption rate, respiration rate, excretion rate, and scope for growth), thermal performance, thermal critical maxima (CTmax), acclimation response ratio (ARR), Arrhenius activation energy (AAE), temperature coefficient (Q10), warming tolerance (WT), and thermal safety margin (TSM) were evaluated. Ingestion and absorption rates were significantly reduced, whereas respiration and ammonia excretion rates significantly increased in stressful treatments, resulting in a significantly lower scope for growth. A profound impact on thermal performance was also noticed, leading to a downward shift in CTmax value for stress-acclimated treatment. The present results clearly highlighted the detrimental combined effect of global climatic stressors and pollution on the physiological energetics of crabs that might potentially reduce their population and affect coastal aquaculture in forthcoming years.

Lack of detrimental effects of ocean acidification and warming on proximate composition, fitness and energy budget of juvenile Senegalese sole (Solea senegalensis)

Rising levels of atmospheric carbon dioxide (CO2) are driving ocean warming and acidification, which may negatively affect the nutritional quality and physiological performance of commercially important fish species. Thus, this study aimed to evaluate the effects of ocean acidification (OA; ΔpH = −0.3 units equivalent to ΔpCO2 ~ +600 μatm) and warming (OW; ΔT = +4 °C) (and combined, OAW) on the proximate composition, fitness and energy budget of juvenile Senegalese sole (Solea senegalensis). After an exposure period of 75 days, growth (G), metabolism (R) and excretion (faecal, F and nitrogenous losses, U) were assessed to calculate the energy intake (C). Biometric and viscera weight data were also registered to determine animal fitness. Overall, the proximate composition and gross energy were not significantly affected by acidification and warming (alone or in combination). Weight gain, maximum and standard metabolic rates (MMR and SMR, respectively), aerobic scope (AS) and C were significantly higher in fish subjected to OA, OW and OAW than in CTR conditions. Furthermore, the highest relative growth rates (RGR), specific growth rates in terms of wet weight (SGRw) and protein (SGRp), as well as feed efficiencies (FE) occurred in fish submitted to OW and OAW. On the other hand, fish exposed to CTR conditions had significantly higher feed conversion ratio (FCR) and ammonia excretion rate (AER) than those exposed to simulated stressors. Regarding energy distribution, the highest fraction was generally allocated to growth (48–63 %), followed by excretion through faeces (36–51 %), respiration (approximately 1 %) and ammonia excretion (0.1–0.2 %) in all treatments. Therefore, ocean warming and acidification can trigger physiological responses in juvenile Senegalese sole, particularly in their energy budget, which can affect the energy flow and allocation of its population. However, and in general, this species seems highly resilient to these predicted ocean climate change stressors.

Research on the molluscicidal activity and molecular mechanisms of arecoline against Pomacea canaliculata

Pomacea canaliculata, as an invasive snail in China, can adversely affect agricultural crop yields, ecological environment, and human health. In this paper, we studied the molluscicidal activity and mechanisms of arecoline against P. canaliculata. The molluscicidal activity tests showed that arecoline exhibits strong toxicity against P. canaliculata, and the LC50 value (72 h) was 1.05 mg/L (15 ± 2 mm shell diameter). Additionally, Molluscicidal toxicity were negatively correlated with the size of snails. Snails (25 ± 2 mm shell diameter) were choosed for mechanisms research and the result of microstructure and biochemistry showed that arecoline (4 mg/L, 20 ℃) had strong toxic effect on the gill, and the main signs were the loss of cilia in the gill filaments. Moreover, arecoline significantly decreased the oxygen consumption rate, ammonia excretion rate and inhibited acetylcholinesterase (AChE). Then, the changes in protein expression were studied by iTRAQ, and 526 downregulated proteins were found. Among these, cilia and flagella-associated 157-like (PcCFP) and rootletin-like (PcRoo) were selected as candidate target proteins through bioinformatics analysis, and then RNA interference (RNAi) was adopted to verify the function of PcCFP and PcRoo. The results showed that after arecoline treated, the mortality and the cilia shedding rate of PcRoo RNAi treated group was significantly lower than control group. The above results indicate that arecoline can bind well with protein PcRoo, and then leads to the drop of gill cilia, affect respiratory metabolism, accelerate its entry into hemolymph, inhibit AChE and finally leads to the death of P. canaliculata.

A simulated toxic assessment of cesium on the blue mussel Mytilus edulis provides evidence for the potential impacts of nuclear wastewater discharge on marine ecosystems

The toxic effects of cesium (Cs) on the blue mussel Mytilus edulis were experimentally investigated to assess the potential environmental consequences of the discharge of nuclear wastewater containing radionuclides. A simulated experimental system of stable cesium (133Cs) was set up to mimic the impacts of radiocesium, and its heavy metal property was emphasized. The mussels were exposed to a concentration gradient of 133Cs for 21 days, followed by another 21-day elimination period. 133Cs exposure resulted in effective bioaccumulation with distinct features of concentration dependence and tissue specificity, and hemolymph, gills and digestive glands were recognized as the most target tissues for accumulation. Although the elimination period was helpful in reducing the accumulated 133Cs, the remaining concentrations of tissues were still significant. 133Cs exposure presented little effect on growth status at the individual level but had distinct interference on feeding and metabolism indicated by the oxygen consumption rate, ammonia-N excretion rate and O:N ratio, simultaneously with the impairment of digestive glands. Regarding hemocytes in the hemolymph, the cell mortality increment, micronucleus promotion, lysosomal membrane stability disruption and phagocytic ability inhibition suggested that the immune function was injured. The cooccurrence of reactive oxygen species overproduction had a close relationship with the observed damages and was thought to be the possible explanation for the immune toxicity. The assay based integrated biomarker response (IBR) presented a good linear relation with the exposure concentrations, suggesting that it was a promising method for assessing the risk of 133Cs. The results indicated that 133Cs exposure damaged M. edulis at the tissue and cell before at the macroscopic individual, evidencing the potentially detrimental impacts of nuclear wastewater discharge on marine ecosystems.

Characterization of two novel ammonia transporters, Hiat1a and Hiat1b, in the teleost model system Danio rerio.

Ammonia excretion in fish excretory epithelia is a complex interplay of multiple membrane transport proteins and mechanisms. Using the model system of zebrafish (Danio rerio) larvae, here we identified three paralogues of a novel ammonia transporter, hippocampus-abundant transcript 1 (DrHiat1), also found in most vertebrates. When functionally expressed in Xenopus laevis oocytes, DrHiat1a and DrHiat1b promoted methylamine uptake in a competitive manner with ammonia. In situ hybridization experiments showed that both transporters were expressed as early as the 4-cell stage in zebrafish embryos and could be identified in most tissues 4days post-fertilization. Larvae experiencing morpholino-mediated knockdown of DrHiat1b exhibited significantly lower whole-body ammonia excretion rates compared with control larvae. Markedly decreased site-specific total ammonia excretion of up to 85% was observed in both the pharyngeal region (site of developing gills) and the yolk sac (region shown to have the highest NH4+ flux). This study is the first to identify DrHiat1b/DrHIAT1 in particular as an important contributor to ammonia excretion in larval zebrafish. Being evolutionarily conserved, these proteins are likely involved in multiple other general ammonia-handling mechanisms, making them worthy candidates for future studies on nitrogen regulation in fishes and across the animal kingdom.

Mechanical and toxicological effects of deep-sea mining sediment plumes on a habitat-forming cold-water octocoral

Deep-sea mining activities are expected to impact deep-sea biota through the generation of sediment plumes that disperse across vast areas of the ocean. Benthic sessile suspension-feeding fauna, such as cold-water corals, may be particularly susceptible to increased suspended sediments. Here, we exposed the cold-water octocoral, Dentomuricea aff. meteor to suspended particles generated during potential mining activities in a four weeks experimental study. Corals were exposed to three experimental treatments: (1) control conditions (no added sediments); (2) suspended polymetallic sulphide (PMS) particles; (3) suspended quartz particles. The two particle treatments were designed to distinguish between potential mechanical and toxicological effects of mining particles. PMS particles were obtained by grinding PMS inactive chimney rocks collected at the hydrothermal vent field Lucky Strike. Both particle types were delivered at a concentration of 25 mg L-1, but achieved suspended concentrations were 2-3 mg L-1 for the PMS and 15-18 mg L-1 for the quartz particles due to the different particle density. Results of the experiment revealed a significant increase in dissolved cobalt, copper and manganese concentrations in the PMS treatment, resulting from the oxidation of sulphides in contact with seawater. Negative effects of PMS exposure included a progressive loss in tissue condition with necrosis and bioaccumulation of copper in coral tissues and skeletons, and death of all coral fragments by the end of the experiment. Physiological changes under PMS exposure, included increased respiration and ammonia excretion rates in corals after 13 days of exposure, indicating physiological stress and potential metabolic exhaustion. Changes in the cellular stress biomarkers and gene expression profiles were more pronounced in corals exposed to quartz particles, suggesting that the mechanical effect of particles although not causing measurable changes in the physiological functions of the coral, can still be detrimental to corals by eliciting cellular stress and immune responses. We hypothesize that the high mortality of corals recorded in the PMS treatment may have resulted from the combined and potentially synergistic mechanical and toxicological effects of the PMS particles. Given the dispersal potential of mining plumes and the highly sensitive nature of octocorals, marine protected areas, buffer areas or non-mining areas may be necessary to protect deep-sea coral communities.

Effects of Temperature on Growth, Molting, Feed Intake, and Energy Metabolism of Individually Cultured Juvenile Mud Crab Scylla paramamosain in the Recirculating Aquaculture System

An eight-week experiment was conducted to investigate the effects of temperature (20, 25, 30, and 35 °C) on growth performance, feed intake, energy metabolism, antioxidant capacity, and the stress response of juvenile Scylla paramamosain in a recirculating aquaculture system. The results showed that the survival rate of the 35 °C group was 80.36 ± 5.92%, significantly lower than that of the other three groups (100%). The high molt frequency of mud crabs was observed in high-temperature groups, accompanied by a higher ecdysone level and ecdysone receptor gene expression but lower molt inhibitory hormone gene expression. However, the molt increment (73.58 ± 2.18%), food intake, and feed conversion efficiency showed a parabolic trend, with the lowest value found in the 35 °C group. Oxygen consumption rate and ammonia excretion rate increased with the increasing temperature, and oxygen-nitrogen ratio, lactic acid, triglyceride, total cholesterol, glucose, and cortisol peaked at 35 °C. Temperature also significantly affected the antioxidant system of S. paramamosain. Crabs in the 25 °C and 30 °C had a significantly higher total antioxidant capacity and lower malondialdehyde compared with the 35 °C group (p < 0.05). Although the high temperature promoted molting, it decreased the feeding rate and growth performance, leading to oxidative stress and functional hypoxia. The quadratic function model demonstrated the optimum temperature for the specific growth rate of juvenile S. paramamosain was 28.5–29.7 °C.

Expression of ABCA3 transporter gene in Tegillarca granosa and its association with cadmium accumulation

Exposure to cadmium (Cd), a heavy metal, can cause strong and toxic side effects. Cd can enter the body of organisms in several ways, leading to various pathological reactions in the body. Tegillarca granosa is a kind of bivalve shellfish favored by people in the coastal areas of China. Bivalve shellfish can easily absorb heavy metal pollutants from water bodies while filter feeding. T. granosa is considered a hyper-accumulator of Cd, and the TgABCA3 gene is highly expressed in individuals with a high content of Cd-exposed blood clam. However, it is unclear whether TgABCA3 is involved in Cd ion transport in blood clam and the molecular mechanism for the mechanism of the Cd-induced responses for maintaining cell homeostasis. In this study, the complete cDNA of the TgABCA3 gene was analyzed to provide insights into the roles of TgABCA3 in resistance against Cd in blood clam. The complete sequence of TgABCA3 showed high identity to that of TgABCA3 from other bivalves and contained some classical motifs of ATP-binding cassette transport proteins. TgABCA3 expression in different tissues was measured using real-time quantitative polymerase chain reaction (qRT-PCR) and western blot analysis. The tissue-specific expression showed that TgABCA3 expression was highest in the gill tissue. The TgABCA3 expression in the gill tissue was silenced using the RNA interference technique. After TgABCA3 silencing, the TgABCA3 expression decreased, the Cd content increased, the oxygen consumption and ammonia excretion rates increased, and the ingestion rate decreased. These results showing that the extents of Cd accumulation and resulting toxic effects are related to expression levels and activity of TgABCA3 indicate that TgABCA3 has a protective function against Cd in the clam. This increase in Cd accumulation results in serious damage to the body, leading to the enhancement of its physiological metabolism. Therefore, the findings of the study demonstrated that TgABCA3 can participate in the transport of Cd ions in the blood clam through active transport and play a vital role in Cd detoxification.